title: Myosin light chain 2-based selection of human iPSC-derived early ventricular cardiac myocytes.
abstract: Applications of human induced pluripotent stem cell derived-cardiac myocytes (hiPSC-CMs) would be strengthened by the ability to generate specific cardiac myocyte (CM) lineages. However, purification of lineage-specific hiPSC-CMs is limited by the lack of cell marking techniques. Here, we have developed an iPSC-CM marking system using recombinant adenoviral reporter constructs with atrial- or ventricular-specific myosin light chain-2 (MLC-2) promoters. MLC-2a and MLC-2v selected hiPSC-CMs were purified by fluorescence-activated cell sorting and their biochemical and electrophysiological phenotypes analyzed. We demonstrate that the phenotype of both populations remained stable in culture and they expressed the expected sarcomeric proteins, gap junction proteins and chamber-specific transcription factors. Compared to MLC-2a cells, MLC-2v selected CMs had larger action potential amplitudes and durations. In addition, by immunofluorescence, we showed that MLC-2 isoform expression can be used to enrich hiPSC-CM consistent with early atrial and ventricular myocyte lineages. However, only the ventricular myosin light chain-2 promoter was able to purify a highly homogeneous population of iPSC-CMs. Using this approach, it is now possible to develop ventricular-specific disease models using iPSC-CMs while atrial-specific iPSC-CM cultures may require additional chamber-specific markers.
authors.affiliation: Department of Internal Medicine, Cardiovascular Medicine, Center for Arrhythmia Research, University of Michigan, Ann Arbor, MI 48108, USA; Université Pierre et Marie Curie, Paris, France.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Bizy
authors.forename: Guadalupe
authors.initials: G
authors.lastname: Guerrero-Serna
authors.forename: Bin
authors.initials: B
authors.lastname: Hu
authors.forename: Daniela
authors.initials: D
authors.lastname: Ponce-Balbuena
authors.forename: B Cicero
authors.initials: BC
authors.lastname: Willis
authors.forename: Manuel
authors.initials: M
authors.lastname: Zarzoso
authors.forename: Rafael J
authors.initials: RJ
authors.lastname: Ramirez
authors.forename: Michelle F
authors.initials: MF
authors.lastname: Sener
authors.forename: Lakshmi V
authors.initials: LV
authors.lastname: Mundada
authors.forename: Matthew
authors.initials: M
authors.lastname: Klos
authors.forename: Eric J
authors.initials: EJ
authors.lastname: Devaney
authors.forename: Karen L
authors.initials: KL
authors.lastname: Vikstrom
authors.forename: Todd J
authors.initials: TJ
authors.lastname: Herron
authors.forename: José
authors.initials: J
authors.lastname: Jalife
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D024722:Cardiac Myosins / Q000235:genetics / Q000378:metabolism*; D002454:Cell Differentiation; D019070:Cell Lineage; D002469:Cell Separation / Q000379:methods*; D005434:Flow Cytometry; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006352:Heart Ventricles / Q000166:cytology*; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000166:cytology* / Q000378:metabolism; D018994:Myosin Light Chains / Q000235:genetics / Q000378:metabolism*; D010641:Phenotype; D011401:Promoter Regions, Genetic; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism
chemical_list: D018994:Myosin Light Chains; D020033:Protein Isoforms; C107184:myosin light chain 2; D049452:Green Fluorescent Proteins; D024722:Cardiac Myosins
keywords: AP; Ad; CM; ESC; FACS; GFP; MLC-2ap; MLC-2vp; action potential; adenovirus; cardiac myocyte; embryonic stem cell; fluorescence-activated cell sorting; green fluorescent protein; hiPSC; human induced pluripotent stem cell; myosin light chain-2 atrial promoter; myosin light chain-2 ventricular promoter